: Fetal serum is enriched for low affinity, self-reactive antibodies, otherwise known as "natural" autoantibodies. Although mouse and man share similar patterns of self-reactivity in fetal life, the mechanisms that yield the limited human repertoire differ from those seen in mice. Fetal antibodies in both mouse and man preferentially use a small subset of highly conserved V and V gene segments. However, unlike mouse, control of CDR 3 diversity and length distribution in man involves control of DH and JH gene segment utilization, and differential N-region addition depending on the identity of the DH gene segment. Fetal antigen binding sites are characterized by selection for an overall neutral charge. High affinity self-reactive antibodies, or "pathologic" autoantibodies, and antibodies found in rheumatoid synovium, are enriched for the use of "fetal" VH and V kappa gene segments, but unlike "fetal" CDR 3 regions, these heavy chains commonly contain JH6, rarely contain DHQ52, and have extensive N-region addition - hallmarks of a "mature" repertoire. These "pathologic" antibodies exhibit significantly greater diversity in their CDR 3 regions. Many of these antibodies have either charged or hydrophobic amino acids.The hypothesis that the polyreactivity of the fetal repertoire is the product of genetic control of gene segment rearrangement, followed by a broad-based selection for an antigen binding site of neutral polarity, will be tested. Populations of sorted pre-B and B-cells from fetal liver and bone marrow, adult bone marrow, and other lymphoid organs will be prepared. The patterns of CDR 3 diversity demonstrated in these sorted populations (N-region addition, DH gene segment utilization and reading frame, JH utilization, and length distribution) will be compared. The patterns of V gene utilization will be sampled through analysis of the usage of members of the related VH1 and VH7 families chosen because of their association with specific B- cell subsets. The extent of self-reactivity encoded by the fetal V gene repertoire will be determined through generation of artificial combinatorial libraries containing either the VH3 gene segment VH26 (30pl) or the VH1 gene segment 51p1 with "fetal" versus "adult"-like H chain CDR3s in association with either Humkv325 or V-kappa 4. If the poly- specificity and self-reactivity of the fetal repertoire is the product of evolutionary selection for auto-reactivity, then the majority of in vitro generated antibodies should be multi-reactive with known panels of self- antigens, whereas self-reactivity should be rare in the "adult" CDR 3 antibodies. Conversely, if the self-reactivity of the fetal repertoire is the product of antigen selection, both libraries should lack this multi-reactive property. Finally, the hypothesis that differential patterns of N-region addition between transcripts that contain DHQ52 versus DXP are due to the presence of two sets of B-cell lineages in early fetal life will be tested through the analoges of fetal pro-B and pre-B-cells. These studies are likely to increase our understanding of the molecular events that limit the fetal repertoire and prevent generation of monospecific, high affinity autoantibodies during ontogeny.